This invention relates to a serum antibody assay used as a diagnostic tool to determine if a particular microorganism might be responsible for a disease state. More particularly, this invention relates to the materials and methods of an assay for serum antibody directed against the particular microorganism which is constructed so as to eliminate false positives caused by the cross-reactivity of other antibodies directed against other microorganisms not targeted by the assay and not the subject of diagnosis.
Frequently, specific serum antibodies can serve as an indirect indicator of a specific disease. A patient's immunogenic response to an infection by a microorganism produces specific antibodies which recognize antigenic sites on that microorganism and which have a binding affinity for such antigenic sites. The detection or measurement of these specific antibodies in the patient's serum provides an indirect method to identify the specific microorganism responsible for the disease. The detection or measurement of these specific antibodies serves as a diagnostic tool for the clinician and enables the clinician to direct therapeutic action against the specific microorganism.
Typically, an antibody assay includes a step in which a sample of the patient's serum is combined with antigenic material derived from the suspect microorganism, i.e. the microorganism to which the assay is targeted. The assay result is positive if serum antibody is found to bind to the antigenic material. However, unless precautionary steps are taken, the positive result of an assay can be, in fact, a false positive.
The patient's serum may contain cross-reactive antibody. Cross-reactive antibody is a product of an immunogenic response to a nontarget microorganism, i.e., a microorganism which is different than the microorganism to which the assay is targeted. However, cross-reactive antibody may bind to the antigenic material used in the assay. Both the target and nontarget microorganisms may share certain structural features among their respective antigens. Antibody which is specific for such a nontarget microorganism may also cross-react with antigen derived from the target microorganism. The presence of cross-reactive antibody can cause false positives in an assay which utilizes antigen derived from the target microorganism. Such cross-reactivity results in a reduction of the diagnostic utility of the assay.
The prior art teaches that this problem of cross-reactivity can be overcome by pretreatment of the patient's serum with antigen derived from the nontarget microorganism having structurally similar antigenic sites to the target microorganism. Such pretreatment will neutralize the cross-reactive antibody, if present, by causing the cross-reactive antibody to bind to the antigen derived from the nontarget microorganism. Such pretreatment will thereby prevent the cross-reactive antibody from binding the antigen of the target microorganism utilized by the assay. An example of an antibody assay with pretreatment is given by Grieve (Grieve, R., et al, American Journal of Veterinary Research, vol. 42 (1), pages 66-69 (1981)). Grieve describes an assay for antibody against Dirofilaria immitis in canine serum which requires a pretreatment of canine sera with Toxocara canis antigen to neutralize anit-T. canis antibodies. Thereafter, the pretreated sera may be tested for anti-D. immitis antibodies substantially without further interference. Unfortunately, such antigen pretreatments introduce additional steps into the assay procedure, which steps may be time consuming and laborious and which may provide additional opportunity for the introduction of error into the assay protocol. Also, additional pretreatments must be made for each nontarget microorganism which is known to elicit cross-reactive antibody to the antigen of the target microorganism.
The present invention addresses the inability of existing antibody assays to measure, without pretreatment, the presence of a specific antibody type as an indication of a specific target microorganism responsible for a disease state, without the risk of the assay yielding a false positive identification caused by cross-reactive serum antibody directed against one or more nontarget microorganisms. For example, the present invention addresses the inability of existing serum antibody assays to detect anti-D. immitis antibody in canine serum so as to diagnose a heartworm infection, without pretreatment of the canine serum and without the risk of obtaining a false positive caused by the cross-reactivity of anti-Toxocara canis antibodies in the canine serum directed against an infection by the intestinal roundworm Toxocara canis.